GSM5751611: RRBS_ESCs_siRNA_Usp7_rep2; Mus musculus; Bisulfite-Seq
DNA samples were extracted using the NucleoSpin RNA purification kit and NucleoSpin RNA/DNA Buffer Set for parallel RNA and DNA purification (Macherey-Nagel) according to the manufacturer's instructions. RRBS libraries were prepared as described in Bender et al. (2021), Methods Mol Biol 2214:207-220. Briefly, 100 ng genomic DNA were digested with MspI (Thermo Fisher Scientific), end-repaired and A-tailed with Klenow fragment exo- (Thermo Fisher Scientific) and ligated to methylated adapters with T4 DNA ligase (Thermo Fisher Scientific) in Tango 1X buffer. DNA fragments ranging from 150 to 400 bp were selected by gel excision, purified using the MinElute gel extraction kit (Qiagen) and bisulfite-converted twice with the EpiTect bisulfite kit (Qiagen). The libraries were amplified using the Pfu Turbo Cx hotstart DNA polymerase (Agilent) with 12 PCR cycles and purified using Agencourt AmpureXP beads (Beckman-Coulter).